The overall objective is to elucidate the mechanism for absorption and secretion of salt and water by vertebrate epithelia. Disordered functions of these mechanisms can lead to a number of disease states including diarrhea, glaucoma, excessive salt and water losses in the urine, and bronchopulmonary diseases, including asthma. All of these epithelia appear to actively absorb or secrete anions (mostly chloride). Closer inspection reveals, however, that Cl transport is only secondarily active, being linked to the Na gradient, which is maintained by Na,K-activated ATPase. The membrand site for coupling Na and Cl fluxes is in the brush border (apical, luminal) membrane of absorptive epithelia and in the basolateral (serosal, blood-side) membrane of secretory epithelia. The goal of this application is to examine these coupling mechanisms, which sometimes involve K as well as Na and Cl. These mechanisms will be examined in two absorptive epithelia (flounder intestine and rabbit proximal colon) and one C1-secreting epithelium (bovine trachea). Radioisotope flux measurements, electrophysiologic techniques, rapid stopped-flow spectrofluorometric methods, radio-labelled diuretic binding to membranes, photoaffinity labelling of the transport sites with an azido-modified diuretic and analyses of cell phosphoproteins will be employed to investigate the coupled transport mechanisms and how they are regulated.